This invention relates to hematology systems and methods. More specifically, this invention relates to systems and methods for analyzing blood samples to identifying, classify, and/or quantify white blood cells (WBC) and WBC sub-populations in a sample of blood.
The development of an accurate and efficient hematology assay for analysis of WBCs, including counting and classification, has been a challenge. One reason for the difficulty in developing an accurate and efficient assay is the relatively low concentration of WBCs (approximately 0.1% to 0.2%) amongst total blood cells in a sample. Attempts to engineer advanced methods for analyzing WBCs, and formulating robust WBC reagent(s), have remained one of the top priorities in the area of automated hematology analyzers.
Typically, lysis of red blood cells (RBCs) is required to eliminate interference from RBCs, and concentrate WBCs, before counting and classifying WBCs and WBC sub-populations. More specifically, accurate and efficient WBC analysis requires: (1) complete lysis of RBCs in less than 30 seconds; (2) the breaking of large fragments of RBCs into smaller pieces after lysis; and (3) the preservation of WBCs for accurate counting and proper classification. If the blood sample is “under-lysed,” unlysed RBCs, even in very small concentrations, interfere with WBC counting and differential analysis. Similarly, larger fragments of lysed RBCs can interfere with WBC counting and differential analysis. In practice, it is difficult to separate unlysed RBCs and/or larger fragments of lysed RBCs from lymphocytes (the smallest WBCs). If the blood sample is “over-lysed,” the classification of WBCs may be adversely affected on account of excessive damage to cell membranes of WBCs.
In some instances, the difficulty of WBC analysis may be compounded by the presence of two special types of samples; namely, samples containing lysis-resistant red blood cells (rstRBCs) and samples containing fragile lymphocytes. In the case of samples containing rstRBCs, the WBC count and percentage of lymphocytes are falsely reported “high,” on account of the contribution of particles other than true lymphocytes, consequently posing a risk of improper diagnoses and treatments for patients. In the case of samples containing fragile lymphocytes, damaged lymphocytes may not show their characteristics in a WBC differential analysis. In addition, exposed nuclei of WBCs may be counted as nucleated red blood cells (nRBCs), resulting in a false positive count of nRBCs in certain assays.